Use of Mytilus galloprovincialis and Tapes philippinarum as sentinel organisms for the development of a biosurveillance program in the Pialassa Baiona coastal lagoon (Ravenna, Italy)

The Pialassa Baiona is an intertidal lagoon located north from the town of Ravenna (Italy). It shows a certain degree of contamination caused by past uncontrolled discharges, and we aim at developing a bioassessment program to be used for future monitoring and conservation plans. A battery of six biomarkers (lysosome membrane stability, lipofuscin and neutral lipid accumulation, hsp70 expression, metallothionein levels and acetylcholinesterase (AChE) activity) was investigated in Mytilus galloprovincialis transplanted for 30 days in different sites. Tapes philippinarum are widely distributed within the lagoon, and we explored the possibility to use resident clams for future biosurveillance programs. Adenylyl cyclase (ACase) activity was also assessed in tissues of both bivalves as a future possible biomarker. Taken together, our results indicate that mussels are suitable sentinel organisms also within such a peculiar environment; alteration of four biomarkers was observed in tissues of mussels transplanted in two sites out of the five analyzed, indicating their low water quality. T. philippinarum showed a barely detectable AChE activity and absence of neutral lipids in digestive glands, therefore appeared unsuitable for our purpose, at least using the selected battery of biomarkers. ACase activity was not different among animals collected from the different sites.     

Differences in response of two model estuarine crustaceans after lethal and sublethal exposures to chlorpyrifos

This study assessed the in vitro and in vivo effects of an acetylcholinesterase enzyme inhibitor (chlorpyrifos) in two estuarine crustaceans: grass shrimp (Palaemonetes pugio) and mysid (Americamysis bahia). The differences in response were quantified after lethal and sublethal exposures to chlorpyrifos and in vitro assays with chlorpyrifos-oxon. Results from the in vitro experiments indicated that the target enzyme, acetylcholinesterase (AChE), in the two species was similar in sensitivity to chlorpyrifos inhibition with IC50s of 0.98 nM and 0.89 nM for grass shrimp and mysids, respectively. In vivo experiments showed that mysids were significantly more sensitive to chlorpyrifos-induced AChE inhibition after 24 h of exposure. The in vivo EC50s for AChE inhibition were 1.23 μg L^?1 for grass shrimp and 0.027 μg L^?1 for mysids.

Median lethal concentrations (24h LC50 values) were 1.06 μg L^?1 for grass shrimp and 0.068 μg L^?1 for mysids. The results suggest that differences in the response of these two crustaceans are likely related to differences in uptake and metabolism rather than target site sensitivity.     

Biamnial elevated alpha-fetoprotein and positive acetylcholinesterase in twins,one with anencephaly

Anencephaly in twin B was accompanied by elevated amniotic fluid alpha-fetoprotein (AFP) and a positive acetylcholinesterase (AChE) band on gel electrophoresis in both twin sacs, although twin A was normal. AChE results did not help distinguish the false positive AFP in this set of twins, implying that AChE may diffuse transamniotically as has been previously postulated for AFP. In light of the low concordance rate for neural tube defects in twins, patient counselling in this situation must include the information that AFP and AChE may be falsely elevated in normal twin when the other twin has a neural tube defect.     

False-positive amniotic fluid acetylcholinesterase analysis in the third trimester

Thirty-two third-trimester amniotic fluid samples were studied according to the indication for amniocentesis, result of acetylcholinesterase (AChE) analysis, and outcome, in order to address the issue of the effectiveness of AChE testing late in gestation. The results indicate that third-trimester AChE analysis is less effective than second trimester in distinguishing open neural tube defects (ONTDs) and ventral wall defects (VWDs) from other abnormalities. False-positive results occurred in cases of isolated hydrocephaly (four of seven cases), polyhydramnios, and intrauterine growth retardation (IUGR). Caution is recommended in interpreting third-trimester AChE tests, particularly when neither an ONTD nor a VWD is observed by ultrasound.     

Amniotic fluid acetylcholinesterase measurement in the prenatal diagnosis of open neural tube defects. Second report of the collaborative acetylcholinesterase study

Seventeen centres from Australia, Britain, France, and the United States collaborated in a study to compare amniotic fluid acetylcholinesterase (AChE) determination by gel electrophoresis and amniotic fluid alpha-fetoprotein (AFP) measurement as diagnostic tests for open neural tube defects. The study was based on 32 642 women with singleton pregnancies (including 428 with open spina bifida and 238 with anencephaly) who had an amniocentesis at 13–24 weeks' gestation. The AChE test yielded a detection rate for open spina bifida of 99 per cent (95 per cent confidence interval 98–100 per cent), 98 per cent for anencephaly (95 per cent confidence interval 96–100 per cent), and a false-positive rate of 0.34 per cent (95 per cent confidence interval 0.28–0.40 per cent) excluding miscarriages, intrauterine death, and serious fetal abnormalities. The false-positive rate was 0.30 per cent among the 13 centres that used a specific AChE inhibitor in the test. Comparable rates for the AFP test were less favourable. (For example, the open spina bifida detection rate was 90 per cent and the false-positive rate was 0.46 per cent using the cut-off levels specified in the U.K. Collaborative AFP Study.) The AChE false-positive rate was lower in samples that were not bloodstained (0.16 per cent) than in those that were (2.4 per cent). It was higher in women who had an amniocentesis on account of a raised maternal serum AFP level (0.56 per cent) than in those who had one for other reasons (0.29 per cent). The best results were obtained by a combination of the two tests, an effective and economical policy being to perform the AFP measurement on all amniotic fluid samples and an AChE test on samples with AFP levels greater than or equal to 2.0 multiples of the normal median (about 5 per cent of all samples). Using this policy, the open spina bifida detection rate was 96 per cent and the false-positive rate was 0.14 per cent (0.06 per cent for samples that were not bloodstained and 1.2 per cent for those that were; 0.40 per cent for women with raised serum AFP levels and 0.09 per cent for other women). This policy offers a useful improvement to the prenatal diagnosis of open spina bifida.     

Dioxin induces expression of hsa-miR-146b-5p in human neuroblastoma cells

Dioxin can cause a series of neural toxicological effects. Micro RNAs(mi Rs) play important roles in regulating nervous system function and mediating cellular responses to environmental pollutants, such as dioxin. Hsa-mi R-146 b-5 p appears to be involved in neurodegenerative diseases and brain tumors. However, little is known about effects of dioxin on the expression of hsa-mi R-146 b-5 p. We found that the hsa-mi R-146 b-5 p expression and its promoter activity were significantly increased in dioxin treated SK-N-SH cells, a human-derived neuroblastoma cell line. Potential roles of hsa-mi R-146 b-5 p in mediating neural toxicological effects of dioxin may be due to the regulation of certain target genes. We further confirmed that hsa-mi R-146 b-5 p significantly suppressed acetylcholinesterase(ACh E) activity and targeted the3′-untranslated region of the ACh E T subunit, which has been down-regulated in dioxin treated SK-N-SH cells. Functional bioinformatic analysis showed that the known and predicted target genes of hsa-mi R-146 b-5 p were involved in some brain functions or cyto-toxicities related to known dioxin effects, including synapse transmission, in which ACh E may serve as a responsive gene for mediating the effect.     

Sublethal Effects of Monocrotophos on Locomotor Behavior and Gill Architecture of the Mosquito Fish,Gambusia affinis

Subacute studies of monocrotophos [Dimethyl (E)-1-methyl-2-(methyl-carbamoyl) vinyl phosphate] on mosquito fish, Gambusia affinis, were carried out in vivo for 24 days to assess the locomotor behavior, structural integrity of gill, and targeted enzyme acetylcholinesterase (AChE, EC: 3.1.1.7) interactions. Monocrotophos (MCP) can be rated as moderately toxic to G. affinis, with a median lethal concentration (LC₅₀) of 20.49 ± 2.45 mgL^?1. The fish exposed to sublethal concentration of LC₁₀ (7.74 mgL^?1) were under stress and altered their locomotor behavior, such as distance traveled per unit time (m min^?1) and swimming speed (cm sec^?1) with respect to the length of exposure. Inhibition in the activity of brain AChE and deformities in the primary and secondary lamellae of gill may have resulted in failure of exchange of gases. The maximum inhibition of 95% of AChE activity was observed on days 20 and 24.

Morphological aberrations in the gills were also studied during exposure to the sublethal concentration of monocrotophos for a period ranging from 8 to 24 days. The extent of damage in gill was dependent on the duration of exposure. The findings revealed that inhibition in brain AChE activity and structural alteration in gill were responsible for altering the locomotor behavior of exposed fish.     

In vitro brain acetylcholinesterase response among three inbred strains of mice to monocrotophos

Abstract

The strain differences in the neurotoxic potential of monocrotophos (HCP) were assessed by determining the inhibition of brain acetylcholinesterase (AChE) in BALB/cAnN, DBA/2J and C57BL/6J in vitro. MCP being a competitive inhibitor for AChE, alters the Km values widely among these inbred strains. Comparatively least alterations in Km were found in BALB/cAnN and maximum in DBA/2J. Based on the Ki values DBA/2J was found to be the most sensitive strain to HCP inhibition followed by C57BL/6J and BALB/cAnN.     

Amniotic fluid acetylcholinesterase in the prenatal diagnosis of open neural tube defects and abdominal wall defects: A comparison of gel electrophoresis and a monoclonal antibody immunoassay

Stored amniotic fluid samples collected in Oxford and East Birmingham as part of the Collaborative Acetylcholinesterase Study were assayed for the presence of acetylcholinesterase (AChE) using a monoclonal antibody (4F19) enzyme antigen immunoassay. These results were compared with the results of a gel AChE which had been performed earlier. A total of 5689 samples from singleton pregnancies were analysed (including 36 with anencephaly, 77 with open spina bifida and 17 with anterior abdominal wall defects). The gel test yielded detection rates of 97% for anencephaly, 99% for open spina bifida and 94% for abdominal wall defects; the false positive rate (excluding pregnancies associated with serious abnormalities, miscarriages and intrauterine deaths) was 0·24%. The monoclonal test yielded similar results; using appropriate cut-off values to allow for differences in acetylcholinesterase levels in blood stained and clear samples, a similar false-positive rate of 0·22% was associated with detection rates of 97%, 95 % and 71 % respectively for the three types of defect. Although the detection rates and the false positive rate were slightly higher for the gel test, a result that might be explained by a decrease in AChE activity caused by storage of the samples, the monoclonal test has the advantages of requiring less interpretative expertise, it can be performed on a larger number of samples a day and it is not affected by contamination with fetal calf serum.